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Chemistry

Investigations on the Ga(III) Complex of EOB-DTPA and Its 68Ga Radiolabeled Analogue

Published: August 17th, 2016

DOI:

10.3791/54334

1Clinic of Nuclear Medicine, University Hospital Jena, 2Friedrich Schiller University

A procedure for the isolation of EOB-DTPA and subsequent complexation with natural Ga(III) and 68Ga is presented herein, as well as a thorough analysis of all compounds and investigations on labeling efficiency, in vitro stability and the n-octanol/water distribution coefficient of the radiolabeled complex.

We demonstrate a method for the isolation of EOB-DTPA (3,6,9-triaza-3,6,9-tris(carboxymethyl)-4-(ethoxybenzyl)-undecanedioic acid) from its Gd(III) complex and protocols for the preparation of its novel non-radioactive, i.e., natural Ga(III) as well as radioactive 68Ga complex. The ligand as well as the Ga(III) complex were characterized by nuclear magnetic resonance (NMR) spectroscopy, mass spectrometry and elemental analysis. 68Ga was obtained by a standard elution method from a 68Ge/68Ga generator. Experiments to evaluate the 68Ga-labeling efficiency of EOB-DTPA at pH 3.8–4.0 were performed. Established analysis techniques radio TLC (thin layer chromatography) and radio HPLC (high performance liquid chromatography) were used to determine the radiochemical purity of the tracer. As a first investigation of the 68Ga tracers' lipophilicity the n-octanol/water distribution coefficient of 68Ga species present in a pH 7.4 solution was determined by an extraction method. In vitro stability measurements of the tracer in various media at physiological pH were performed, revealing different rates of decomposition.

Gadoxetic acid, a common name for the Gd(III) complex of the ligand EOB-DTPA1, is a frequently used contrast agent in hepatobiliary magnetic resonance imaging (MRI).2,3 Due to its specific uptake by liver hepatocytes and high percentage of hepatobiliary excretion it enables the localization of focal lesions and hepatic tumors.2-5 However, certain limitations of the MRI technique (e.g., toxicity of the contrast agents, limited applicability in patients with claustrophobia or metal implants) call for an alternative diagnostic tool.

Positron emission tomography (PET) is a molecular imaging method, wher....

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1. Preparation of EOB-DTPA and Ga[EOB-DTPA]

Caution: Please consult all relevant material safety data sheets (MSDS) of the used organic solvents, acids and alkalines before use. Perform all steps in a fume hood and use personal protective equipment (safety glasses, gloves, lab coat).

  1. Isolation of EOB-DTPA from gadoxetic acid
    1. Put 3 ml of 0.25 M gadoxetic acid injectable solution into a flask. Add 500 mg (5.6 mmol) of oxalic acid to the stirred solution.
    2. After stir.......

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The ligand EOB-DTPA and the non-radioactive Ga(III) complex were analyzed via 1H and 13C{1H} NMR spectroscopy, mass spectrometry and elemental analysis. The results listed in Table 1 and depicted in Figures 1-6 verify the purity of the substances.

Elution of the 68Ge/68Ga generator yielded solutions of 400-600 MBq 68Ga. T.......

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EOB-DTPA is accessible through a multi-step synthesis33 but may just as well be isolated from available contrast agents containing gadoxetic acid. For this purpose, the central Gd(III) ion can be precipitated with an excess of oxalic acid. After removing Gd(III) oxalate and oxalic acid the ligand can be isolated by precipitation in cold water at pH 1.5. However, in order to enhance yields column chromatography of the filtrate can be performed instead or as a follow-up procedure. Either method yields the analyt.......

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The authors have no acknowledgements.

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Name Company Catalog Number Comments
primovist Bayer - 0.25 M
gallium(III) chloride Sigma-Aldrich Co. 450898
water (deionized) - -  tap water deionizing equipment by Auma-Tec GmbH
hydrochloric acid 12 M VWR 20252.29
sodium hydroxide Polskie Odczynniki Chemiczne S.A. 810925429
oxalic acid Sigma-Aldrich Co. 75688
ethyl acetate Brenntag GmbH 10010447
silica gel Merck KGaA 1.10832.9025 Geduran Si 60 0,063-0,2 mm
TLC silica gel 60 F254 Merck KGaA 1.16834.0001
methanol VWR 20903.55
ethanol Brenntag GmbH 10018366
eiethylether VWR 23807.468 stored over KOH plates
ammonia solution (25 %) VWR 1133.1
pH electrode VWR 662-1657
stirring and heating unit Heidolph 505-20000-00
pump Ilmvac GmbH 322002
frit - custom design
NMR spectrometer Bruker Coorporation - Ultra Shield 400
mass spectrometer Thermo Fisher Scientific Inc. -
elemental analyser Hekatech GmbH Analysentechnik - EuroVector EA 3000 CHNS
deuterated water D2O euriso-top D214 99,90 % D
Name Company Catalog Number Comments
Material/Equipment required for labeling procedures
68Ge/68Ga generator ITG Isotope Technologies Garching GmbH A150
pump and dispenser system Scintomics GmbH - Variosystem
hydrochloric acid 30 % (suprapur) Merck KGaA 1.00318.1000
water (ultrapur) Merck KGaA 1.01262.1000
sodium chloride (suprapur) Merck KGaA 1.06406.0500
sodium acetate (suprapur) Merck KGaA 1.06264.0050
glacial acetic acid (suprapur) Merck KGaA 1.00066.0250
sodium citrate dihydrate VEB Laborchemie Apolda 10782 >98.5%
PS-H+ Cartridge (S) Macherey-Nagel 731867 Chromafix
apo-Transferrin Sigma-Aldrich Co. T2036
PBS  buffer (tablets) Sigma-Aldrich Co. 79382
human serum Sigma-Aldrich Co. H4522 from human male AB plasma
flasks, columns etc. custom design
pH electrode Knick Elektronische Messgeräte GmbH & Co. KG 765-Set
binary pump (HPLC) Hewlett-Packard G1312A (HP 1100)
UV Vis detector (HPLC) Hewlett-Packard G1315A (HP 1100)
radioactive detector (HPLC) EGRC Berthold
HPLC C-18-PFP column Advanced Chromatography Technologies Ltd. ACE-1110-1503/A100528
HPLC glass vials GTG Glastechnik Graefenroda GmbH 8004-HP-H/i3µ
pipette Eppendorf -
plastic vials Sarstedt AG & Co. 6542.007
plastic vials Greiner Bio-One International GmbH 717201
activimeter MED Nuklear-Medizintechnik Dresden GmbH - Isomed 2010
tweezers custom design
incubator Heraeus Instruments GmbH 51008815
vortex mixer Fisons - Whirlimixer
centrifuge Heraeus Instruments GmbH 75003360
gamma well counter MED Nuklear-Medizintechnik Dresden GmbH - Isomed 2100
water for chromatography Merck KGaA 1.15333.2500
acetonitrile for chromatography Merck KGaA 1.00030.2500
trifluoroacetic acid Sigma-Aldrich 91707
TLC radioactivity scanner raytest Isotopenmessgeräte GmbH B00003875 equipped with beta plastic detector

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